Compounds and compositions for delivering active agents

ABSTRACT

The present invention provides delivery agent compounds, compositions containing delivery agent compounds and an active agent and methods for delivering active agents, such as biologically or chemically active agents.

CROSS-REFERENCE TO PRIOR APPLICATIONS

This is a U.S. National Phase Application under U.S.C. §371 ofInternational Patent Application No. PCT/US2007/077100 filed Aug. 29,2007, which claims the benefit of U.S. Provisional Application No.60/841,723, filed Aug. 31, 2006. The International Applicationre-published in English on Aug. 21, 2008 as WO2008/027958 under Article21(2).

PRIORITY

The present invention claims the priority of U.S. Application No.60/841,723, filed on 31 Aug. 2006.

FIELD OF THE INVENTION

The present invention relates to compounds, compositions and methods fordelivering active agents, such as biologically or chemically activeagents.

BACKGROUND OF THE INVENTION

Conventional means for delivering active agents are often severelylimited by biological, chemical and physical barriers. Typically, thesebarriers are imposed by the environment through which delivery occurs,the environment of the target for delivery, and/or the target itself.Biologically and chemically active agents are particularly vulnerable tosuch barriers.

In the delivery to animals of biologically active and chemically activepharmacological and therapeutic agents, barriers are also imposed by thebody. Examples of physical barriers are the skin, lipid bi-layers andvarious organ membranes that are relatively impermeable to certainactive agents but must be traversed before reaching a target, such asthe circulatory system. Chemical barriers include, but are not limitedto, pH variations in the gastrointestinal (GI) tract and degradingenzymes.

These barriers are of particular significance in the design of oraldelivery systems. Oral delivery of many biologically or chemicallyactive agents would be the route of choice for administration to animalsif not for biological, chemical, and physical barriers. Among thenumerous agents which are not typically amenable to oral administrationare biologically or chemically active peptides, such as calcitonin andinsulin; polysaccharides, and in particular mucopolysaccharidesincluding, but not limited to, heparin; heparinoids; antibiotics; andother organic substances. These agents may be rapidly renderedineffective or destroyed in the gastro-intestinal tract by acidhydrolysis, enzymes, and the like. In addition, the size and structureof macromolecular drugs may prohibit absorption.

Earlier methods for orally administering vulnerable pharmacologicalagents have relied on the co-administration of adjuvants (e.g.,resorcinols and non-ionic surfactants such as polyoxyethylene oleylether and n-hexadecylpolyethylene ether) to increase artificially thepermeability of the intestinal walls, as well as the co-administrationof enzymatic inhibitors (e.g., pancreatic trypsin ihibitors,diisopropylfluorophosphate (DFF) and trasylol) to inhibit enzymaticdegradation. Liposomes have also been described as drug delivery systemsfor insulin and heparin. However, broad spectrum use of such drugdelivery systems is precluded because: (1) the systems require toxicamounts of adjuvants or inhibitors; (2) suitable low molecular weightcargos, i.e. active agents, are not available; (3) the systems exhibitpoor stability and inadequate shelf life; (4) the systems are difficultto manufacture; (5) the systems fail to protect the active agent(cargo); (6) the systems adversely alter the active agent; or (7) thesystems fail to allow or promote absorption of the active agent.

Proteinoid microspheres have been used to deliver pharmaceuticals. See,for example, U.S. Pat. Nos. 5,401,516; 5,443,841; and Re. 35,862. Inaddition, certain modified amino acids have been used to deliverpharmaceuticals. See, for example, U.S. Pat. Nos. 5,629,020; 5,643,957;5,766,633; 5,776,888; and 5,866,536.

More recently, a polymer has been conjugated to a modified amino acid ora derivative thereof via a linkage group to provide for polymericdelivery agents. The modified polymer may be any polymer, but preferredpolymers include, but are not limited to, polyethylene glycol (PEG), andderivatives thereof See, for example, International Patent PublicationNo. WO 00/40203.

However, there is still a need for simple, inexpensive delivery systemswhich are easily prepared and which can deliver a broad range of activeagents by various routes.

SUMMARY OF THE INVENTION

The present invention relates to compounds (hereafter referred to as“delivery agent compounds”) that facilitate the delivery of activeagents. The delivery agent compound of the present invention may beselected from the group consisting of:

and pharmaceutically acceptable salts thereof.

Mixtures of these delivery agent compounds may also be used.

The invention also provides a composition (e.g., pharmaceuticalcompositions) comprising at least one of the delivery agent compounds,and at least one active agent. These compositions deliver active agentsto biological systems with increased or improved bioavailability of theactive agent compared to administration of the active agent without thedelivery agent compound.

Also provided is a dosage unit form comprising the composition of thepresent invention. The dosage unit form may be in the form of a liquidor a solid, such as a tablet, capsule or particle, including a powder orsachet.

Another embodiment is a method for administering an active agent to ananimal, particularly an animal in need of the active agent, byadministering a composition comprising at least one of the deliveryagent compounds and the active agent to the animal. Preferred routes ofadministration include the oral and intracolonic routes.

Yet another embodiment is a method of treating a disease or forachieving a desired physiological effect in an animal by administeringan effective amount of the composition of the present invention.

Yet another embodiment is a method of preparing a composition of thepresent invention by mixing at least one delivery agent compound of theformulas above, and at least one active agent.

BRIEF DESCRIPTION OF THE DRAWINGS

FIGS. 1-6 are graphs of plasma argatroban concentrations in male,Sprague-Dawley rats versus time after intravenous and oraladministration of argatroban without a delivery agent, and oraladministration of argatroban with delivery agent compounds of thepresent invention. FIG. 7 contains the references of compounds used inFIGS. 1-6.

DETAILED DESCRIPTION OF THE INVENTION

Delivery Agent Compounds

The delivery agent compound (e.g. 4-(2,5-dimethylphenoxy)butyric acid)may be in the form of its free acid or salts thereof such aspharmaceutically acceptable salts. Suitable salts include, but are notlimited to, organic and inorganic salts, for example, ammonium, acetatesalt, citrate salt, halide (preferably hydrochloride), alkali metal(e.g., sodium and potassium), hydroxide, sulfate, nitrate, phosphate,alkoxy, perchlorate, tetrafluoroborate, carboxylate, mesylate, fumerate,malonate, succinate, tartrate, acetate, gluconate, and maleate.Preferred salts include, but are not limited to, sodium, citrate andmesylate salts. The salts may also be solvates, including ethanolsolvates, and hydrates.

Salts of the delivery agent compounds of the present invention may beprepared by methods known in the art. For example, citrate salts may beprepared in ethanol, toluene and citric acid.

The delivery agent compound may be purified by recrystallization or byfractionation on one or more solid chromatographic supports, alone orlinked in tandem. Suitable recrystallization solvent systems include,but are not limited to, ethanol, water, heptan, ethyl acetate,acetonitrile, acetone, methanol, and tetrahydrofuran (THF) and mixturesthereof Fractionation may be performed on a suitable chromatographicsupport such as alumina, using methanol/n-propanol mixtures as themobile phase; reverse phase chromatography using trifluoroaceticacid/acetonitrile mixtures as the mobile phase; and ion exchangechromatography using water or an appropriate buffer as the mobile phase.When anion exchange chromatography is performed, preferably a 0-500 mMsodium chloride gradient is employed.

The delivery agent may contain a polymer conjugated to it by a linkagegroup selected from the group consisting of —NHC(O)NH—, —C(O)NH—,—NHC(O), —OOC—, —COO—, —NHC(O)O—, —OC(O)NH—, —CH₂NH—NHCH₇—, —CH₂NHC(O)—,—OC(O)NHCH₂—, —CH₂NHCOCH₂O—, —OCH₂C(O)NHCH₂—, —NHC(O)OH₂O—,—OCH₂C(O)NH—, —NH—, —O—, and carbon-carbon bond. According to oneembodiment, the polymeric delivery agent is not a polypeptide orpolyamino acid. Such polymer-delivery agent conjugates and methods forpreparing them are described in International Published Application No.WO 00/40203, which is hereby incorporated by reference. The polymer maybe any polymer including, but not limited to, alternating copolymers,block copolymers and random copolymers, which are safe for use inmammals. Preferred polymers include, but are not limited to,polyethylene; polyacrylates; polymethacrylates; poly(oxyethylene);poly(propylene); polypropylene glycol; polyethylene glycol (PEG); andderivatives thereof and combinations thereof. The molecular weight ofthe polymer typically ranges from about 100 to about 200,000 daltons.The molecular weight of the polymer preferably ranges from about 200 toabout 10,000 daltons. In one embodiment, the molecular weight of thepolymer ranges from about 200 to about 600 daltons and more preferablyranges from about 300 to about 550 daltons.

Active Agents

Active agents suitable for use in the present invention includebiologically active agents and chemically active agents, including, butnot limited to, pesticides, pharmacological agents, and therapeuticagents. Suitable active agents include those that are rendered lesseffective, ineffective or are destroyed in the gastro-intestinal tractby acid hydrolysis, enzymes and the like. Also included as suitableactive agents are those macromolecular agents whose physiochemicalcharacteristics, such as, size, structure or charge, prohibit or impedeabsorption when dosed orally.

For example, biologically or chemically active agents suitable for usein the present invention include, but are not limited to, proteins;polypeptides; peptides; hormones; polysaccharides, and particularlymixtures of muco-polysaccharides; carbohydrates; lipids; small polarorganic molecules (i.e. polar organic molecules having a molecularweight of 500 daltons or less); other organic compounds; andparticularly compounds which by themselves do not pass (or which passonly a fraction of the administered dose) through the gastro-intestinalmucosa and/or are susceptible to chemical cleavage by acids and enzymesin the gastrointestinal tract; or any combination thereof.

Further examples include, but are not limited to, the following,including synthetic, natural or recombinant sources thereof: growthhormones, including human growth hormones (hGH), recombinant humangrowth hormones (rhGH), bovine growth hormones, and porcine growthhormones; growth hormone releasing hormones; growth hormone releasingfactor, interferons, including α (e.g., interferon alfacon-1 (availableas Infergen® from InterMune, Inc. of Brisbane, Calif.)), β and γ;interleukin-1; interleukin-2; insulin, including porcine, bovine, human,and human recombinant, optionally having counter ions including zinc,sodium, calcium and ammonium; insulin-like growth factor, includingIGF-1; heparin, including unfractionated heparin, heparinoids,dermatans, chondroitins, low molecular weight heparin, very lowmolecular weight heparin and ultra low molecular weight heparin;calcitonin, including salmon, eel, porcine and human; erythropoietin;atrial naturetic factor; antigens; monoclonal antibodies; somatostatin;protease inhibitors; adrenocorticotropin, gonadotropin releasinghormone; oxytocin; leutinizing-hormone-releasing-hormone; folliclestimulating hormone; glucocerebrosidase; thrombopoietin; filgrastitn;prostaglandins; cyclosporin; vasopressin; cromolyn sodium (sodium ordisodium chromoglycate); vancomycin; desferrioxamine (DFO);bisphosphonates, including alendronate, tiludronate, etidronate,clodronate, pamidronate, olpadronate, and incadronate; parathyroidhormone (PTH), including its fragments; anti-migraine agents such asBIBN-4096BS and other calcitonin gene-related proteins antagonists;glucagon-like peptide 1 (GLP-1); argatroban; antimicrobials, includingantibiotics, anti-bacterials and anti-fungal agents; vitamins; analogs,fragments, mimetics or polyethylene glycol (PEG)-modified derivatives ofthese compounds; or any combination thereof. Non-limiting examples ofantibiotics include gram-positive acting, bacteriocidal, lipopeptidaland cyclic peptidal antibiotics, such as daptomycin and analogs thereof.

Delivery Systems

The composition of the present invention comprises one or more deliveryagent compounds of the present invention, and one or more active agents.In one embodiment, one or more of the delivery agent compounds, or saltsof these compounds, or poly amino acids or peptides of which thesecompounds or salts form one or more of the units thereof, may be used asa delivery agent by mixing with the active agent prior to administrationto form an administration composition.

The administration compositions may be in the form of a liquid. Thesolution medium may be water (for example, for salmon calcitonin,parathyroid hormone, and erythropoietin), 25% aqueous propylene glycol(for example, for heparin) and phosphate buffer (for example, for rhGH).Other dosing vehicles include polyethylene glycol. Dosing solutions maybe prepared by mixing a solution of the delivery agent compound with asolution of the active agent, just prior to administration.Alternatively, a solution of the delivery agent compound (or activeagent) may be mixed with the solid form of the active agent (or deliveryagent compound). The delivery agent compound and the active agent mayalso be mixed as dry powders. The delivery agent compound and the activeagent can also be admixed during the manufacturing process.

The dosing solutions may optionally contain additives such as phosphatebuffer salts, citric acid, glycols, or other dispersing agents.Stabilizing additives may be incorporated into the solution, preferablyat a concentration ranging between about 0.1 and 20% (w/v).

The administration compositions may also be in the form of a solid, suchas a tablet, capsule or particle, such as a powder or sachet. Soliddosage forms may be prepared by mixing the solid form of the compoundwith the solid form of the active agent. Alternatively, a solid may beobtained from a solution of compound and active agent by methods knownin the art, such as freeze-drying (lyophilization), precipitation,crystallization and solid dispersion.

The administration compositions of the present invention may alsoinclude one or more enzyme inhibitors. Such enzyme inhibitors include,but are not limited to, compounds such as actinonin or epiactinonin andderivatives thereof. Other enzyme inhibitors include, but are notlimited to, aprotinin (Trasylol) and Bowman-Birk inhibitor.

The amount of active agent used in an administration composition of thepresent invention is an amount effective to accomplish the purpose ofthe particular active agent for the target indication. The amount ofactive agent in the compositions typically is a pharmacologically,biologically, therapeutically, or chemically effective amount. However,the amount can be less than that amount when the composition is used ina dosage unit form because the dosage unit form may contain a pluralityof delivery agent compound/active agent compositions or may contain adivided pharmacologically, biologically, therapeutically, or chemicallyeffective amount. The total effective amount can then be administered incumulative units containing, in total, an effective amount of the activeagent.

The total amount of active agent to be used can be determined by methodsknown to those skilled in the art. However, because the compositions ofthe invention may deliver active agents more efficiently thancompositions containing the active agent alone, lower amounts ofbiologically or chemically active agents than those used in prior dosageunit forms or delivery systems can be administered to the subject, whilestill achieving the same blood levels and/or therapeutic effects.

The presently disclosed delivery agent compounds facilitate the deliveryof biologically and chemically active agents, particularly in oral,intranasal, sublingual, intraduodenal, subcutaneous, buccal,intracolonic, rectal, vaginal, mucosal, pulmonary, transdermal,intradermal, parenteral, intravenous, intramuscular and ocular systems,as well as traversing the blood-brain barrier.

Dosage unit forms can also include any one or combination of excipients,diluents, disintegrants, lubricants, plasticizers, colorants,flavorants, taste-masking agents, sugars, sweeteners, salts, and dosingvehicles, including, but not limited to, water, 1,2-propane diol,ethanol, olive oil, or any combination thereof.

The compounds and compositions of the subject invention are useful foradministering biologically or chemically active agents to animals,including but not limited to birds such as chickens; mammals, such asrodents, cows, pigs, dogs, cats, primates, and particularly humans; andinsects.

The system is particularly advantageous for delivering chemically orbiologically active agents that would otherwise be destroyed or renderedless effective by conditions encountered before the active agent reachesits target zone (i.e. the area in which the active agent of the deliverycomposition is to be released) and within the body of the animal towhich they are administered. Particularly, the compounds andcompositions of the present invention are useful for orallyadministering active agents, especially those that are not ordinarilyorally deliverable, or those for which improved delivery is desired.

The compositions comprising the delivery agent compounds and activeagents have utility in the delivery of active agents to selectedbiological systems and in an increased or improved bioavailability ofthe active agent compared to administration of the active agent withoutthe delivery agent. Delivery can be improved by delivering more activeagent over a period of time, or in delivering the active agent in aparticular time period (such as to effect quicker or delayed delivery),or in delivering the active agent at a specific time, or over a periodof time (such as sustained delivery).

Another embodiment of the present invention is a method for thetreatment or prevention of a disease or for achieving a desiredphysiological effect, such as any one of the diseases or conditionslisted in the table below, in an animal by administering the compositionof the present invention. Preferably, an effective amount of thecomposition for the treatment or prevention of the desired disease orfor achieving the desired physiological effect is administered. Specificindications for active agents can be found in the The Physicians' DeskReference (58^(th) Ed., 2004, Medical Economics Company, Inc., Montvale,N.J.), and Fauci, A S, et. al., Harrison's Principles of InternalMedicine (14^(th) Ed., 1998, McGraw-Hill Health Professions Division,New York. Both of these references are herein incorporated by referencein their entirety. The active agents in the table below include theiranalogs, fragments, mimetics, and polyethylene glycol-modifiedderivatives (e.g., the PEGylated derivative of granulocyte colonystimulating factor sold as Neulasta®).

Active Agent Disease and Physiological Effect Growth hormones (includinghuman Growth disorders recombinant growth hormone and growth- hormonereleasing factors and its analogs) Interferons, including α, β and γViral infection, including chronic cancer, hepatitis, and multiplesclerosis Interleukins (e.g. Interleukin-1; interleukin-2) Viralinfection; cancer; cell mediated immunity; and transplant rejection;Insulin; Insulin-like growth factor IGF-1 Diabetes Immune Globulins,such as IVIg smallpox, rabies, and diphtheria, Alzheimer's Disease;Primary immunodeficiencies; Acute Guillain-Barre syndrome; Chronicidiopathic demyelinating polyneuropathy (CIDP); Myasthenia gravis,polymyositis, and dermatomyositis; neonatal immune thrombocytopenia,heparin-induced thrombocytopenia, and antiphospholipid antibodysyndrome: Posttransfilsion purpura. Heparin Treatment and Prevention ofThrombosis, including (Deep Vein Thrombosis); prevention of bloodcoagulation Calcitonin Osteoporosis; diseases of the bone; bone pain;analgesic (including pain associated with osteoporosis or cancer)Erythropoietin, Pegylated erythropoietin. Anemia; HIV/HIV-therapyAssociated Anemia; Chemotherapeutically-Induced Anemia Atrial natureticfactor Vasodilation Antigens Infection CPHPC Reduction of amyloiddeposits and systemic amyloidoisis often (but not always) in connectionwith Alzheimer's disease, Type II diabetes, and other amyloid-baseddiseases Monoclonal antibodies To prevent graft rejection; cancer; usedin assays to detect diseases Somatostatin/octreotide Bleeding ulcer;erosive gastritis; variceal bleeding; diarrhea; acromegaly;TSH-secreting pituitary adenomas; secretory pancreatic tumors; carcinoidsyndrome; reduce proptosis/thyroid- associated ophthalmopathy; reducemacular edema/retinopathy Protease inhibitors HIV Infection/AIDSAdrenocorticotropin High cholesterol (to lower cholesterol) Gonadotropinreleasing hormone Ovulatory disfunction (to stimulate ovulation)Oxytocin Labor disfunction (to stimulate contractions)Leutinizing-hormone-releasing-hormone; Regulate reproductive functionLeutinizing Hormone; follicle stimulating hormone GlucocerebrosidaseGaucher disease (to metabolize lipoprotein) ThrombopoietinThrombocytopenia Filgrastim (Granulocyte Colony Stimulating shorten theduration of chemotherapy-induced Factor); GM-CSF, (sargramostim) andtheir neutropenia and thus treat or prevent infection Pegylated forms inchemotherapy patients; Inhibit the growth of or to kill MycobacteriumIntracellular Avium Infection (MAC) RNAi Huntington, Alzheimers, ViralInfections (HIV, Hepatitis A, B or C, RSV), Cancers; MacularDegeneration Prostaglandins Hypertension Cyclosporin Transplantrejection; psoriasis, inflammatory alopecias; Sjogren's syndrome;Keratoconjunctivitis Sicca Vasopressin Nocturnal Enuresis; antidiureticCromolyn sodium; Asthma; allergies Vancomycin Treat or preventantimicrobial-induced infections including, but not limitted tomethacillin-resistant Staphaloccccus aureus and Staph. epidermiditisgallium salts (such as gallium nitrate) Osteoporosis; Paget's disease;Inhibits osteoclasts; Promotes osteoblastic activity, hypercalcemia,including cancer related hypercalcemia, urethral (urinary tract)malignancies; anti-tumors, cancers, including urethral and bladdercancers; lymphoma; malignancies (including bladder cancer); leukemia;management of bone metastases (and associated pain); muliple myeloma,attenuate immune response, including allogenic transplant rejections;disrupt iron metabolism; promote cell migration; wound repair; toattenuate or treat infectious processes of mycobacterium species,including but not limited to mycobacterium tubercolosis, andmycobacterium avium complex Desferrioxamine (DFO) Iron overloadParathyroid hormone (PTH), including its Osteoporosis; fragments.Diseases of the bone Antimicrobials Infection including but not limitedto gram- positive bacterial infection Vitamins Treat and prevent Vitamindeficiencies Bisphosphonates Osteoporosis; Paget's disease; bone tumorsand metastases (and associated pain); Breast cancer; including asadjuvant therapy for early stage breast cancer; management of bonemetastases (and associated pain), including bone metastases associatewith breast cancer, prostate cancer, and lung cancer; Inhibitsosteoclasts; Promotes osteoblastic activity; treat and/or prevent bonemineral density (bmd) loss; multiple myeloma; prevention of bonecomplications related to malignant osteolysis; fibrous dysplasia;pediatric osteogenesis imperfecta; hypercalcemia, urethral (urinarytract) malignancies; reflex sympathetic dystropy synodrome, acute backpain after vertebral crush fracture, chronic inflammatory joint disease,renal bone disease, extrosseous calcifications, analgesic, vitamin Dintoxication, periarticular ossifications BIBN4096BS -(1-Piperidinecarboxamide·N- Anti-migraine; calcitonin gene- relatedpeptide [2-[[5-amino-1-[[4-(4-pyridinyl)-1- antagonistpiperazinyl)carbonyl]pentyl]amino]-1-[(3,5-dibromo-4-hydroxyphenyl)methyl]-2- oxoethyk]-4(1,4-dihydro-2-oxo-3(2H0-quinazolinyl)-·[R-(R*,S*)]-) Glucagon improving glycemic control (e.g.treating hypoglycemia and controlling hypoglycemic reactions), obesity;a diagnostic aid in the radiogical examination of the stomach, duodenum,small bowel and colon; Treat acute poisoning With Cardiovascular Agentsincluding, but not limited to, calcium channel blockers, beta blockersGLP-1, Exendin - 3, Exendin -4, Obestatin Diabetes; improving glycemiccontrol (e.g. treating hypoglycemia and controlling hypoglycemicreactions), obesity dipeptidyl peptidase IV (DPP-4) inhibitors Diabetes;improving glycemic control (e.g. treating hypoglycemia), obesityacyclovir Used to treat herpes infections of the skin, lip and genitals;herpes zoster (shingles); and chickenpox HIV Entry Inhibitors (e.g.Fuzeon) Inhibit entry of HIV into host cells Sumatriptin, almotriptan,naratriptan, anti-migraine serotonin agonists rizatriptan, frovatriptanand eletriptan (piperidinyloxy)phenyl, (piperidinyloxy)pyridinyl,(piperidinylsulfanyl)phenyl and (piperidinylsulfanyl)pyridinyl compoundsNeuraminidase inhibitors: peramivir, Antivirals zanamivir, oseltamivir,BCX-1898, BCX-1827, BCX-1989, BCX 1923, BCX 1827 and A315675; M2inhibitors: amantadine, rimantadine; Nucleoside/Nucleotide ReverseTranscriptase Inhibitors, Non-nucleoside Reverse TranscriptaseInhibitors, Protease Inhibitors, Fusion inhibitors: thiovir,thiophosphonoformate, foscarnet, enfuviritide, zidovudine, didanosine,zalcitabine, stavudine, lamivudine, emtricitabine, abacavir,azidothymidine, tenofovir disoproxil, delavridine, efavirenz,nevirapine, ritonavir, nelfinavir mesylate, saquinvir mesylate,indinavir sulfate, amprenavir, lopinavir, lopinavir, fosamprenavircalcium, atazanavir sulfate Peptide YY (PYY) and PYY-like Peptides (e.g.Obesity, Diabetes, Eating Disorders, Insulin- PYY[3-36]) ResistanceSyndromes

For example, one embodiment of the present invention is a method fortreating a patient having or susceptible to diabetes by administeringinsulin in a pharmaceutical formulation of the present invention. Otheractive agents, including those set forth in the above table, can be usedin conjunction with the pharmaceutical formulations of the presentinvention.

Following administration, the active agent present in the composition ordosage unit form is taken up into the circulation. The bioavailabilityof the agent can be readily assessed by measuring a knownpharmacological activity in blood, e.g. an increase in blood clottingtime caused by heparin, or a decrease in circulating calcium levelscaused by calcitonin. Alternately, the circulating levels of the activeagent itself can be measured directly.

EXAMPLES

The following examples illustrate the present invention withoutlimitation.

Example 1 Preparation of 4-(2,5-Dimethylphenoxy)butyric acid (Compound1)

2,5-Dimethylphenol (6.1 g, 0.05 mol) and ethyl bromobutyrate weredissolved in 60 ml of N,N-Dimethylacetamide (DMAC). Potassium carbonate(11 g) and potassium iodide 0.5 g (0.003 mol) were added to thissolution. The mixture was stirred at 70-80° C. for 24 hours. Thefiltrate was added to distilled water (200 ml). An oily precipitateformed and was extracted with methylene chloride (100 ml). The organiclayer was washed with 5% sodium carbonate (3×150 ml), dried overanhydrous sodium sulfate, and concentrated to give al oily residue. Theoil was dissolved in ethanol (5 ml). The solution was mixed with 2Nsodium hydroxide (30 ml). The mixture was stirred at 70-80° C. until theester was hydrolyzed (determined by HPLC). The solution was acidifiedwith 3N hydrochloride to pH 1 to form a precipitate. The precipitate wascollected by filtration and dissolved in 4% sodium bicarbonate.Insoluble materials were removed by filtration. The filtrate wasacidified to pH 1 to form a precipitate. The precipitate was collectedby filtration and air-dried. 6.6 g of 4-(2,5-dimethylphenoxy) butyricacid was recovered. Melting point: 63-65° C. Combustion analysis: % C.69.21 (calcuated), 69.04 (found); % H, 7.74 (calculated); 7.49 (found).1H NMR Analysis (d6-DMSO). δ 6.98, d, 1H; 6.71, s, 1H; 6.62, d, 1H;3.94, t, 2H; 2.40, t, 2H; 2.24, s, 3H; 2.09, s, 3H; 1.95, p, 2H.

Example 2 Preparation of 4-(2,5-Dimethylphenoxy)butyric acid (Compound1)

2,5-dimethylphenol (24.5 g, 0.2 mol) was dissolved in 125 ml ofanhydrous ethanol. To the solution was added 75 ml of 21 wt % sodiumethoxide solution in ethanol (0.2 mol). The mixture solution was stirredat room temperature for 10 min. Ethyl 4-bromobutyrate (40 g, 0.205 mol)was then added to the solution. The reaction mixture was refluxed at 80°C. for 24 h. The precipitate was filtered off. The filtrate was thenmixed with a solution of sodium hydroxide (10 g, 0.25 mol) in 150 ml ofwater. The mixture was stirred at 70° C. for about 2 hours, monitoredwith HPLC. It was diluted with 150 ml of water and acidified to pH 1.Oily precipitate formed immediately. It was collected by decanting thesupernatant and washed with water twice (100 ml×2). The oily precipitatesolidified after kept in water at room temperature overnight.

The precipitate was dissolved in 250 ml of water at pH 8-9. Anyinsoluble substance was filtered off. The filtrate was extracted withethyl acetate twice (100 ml×2). The aqueous solution was acidified topH 1. Precipitate formed immediately as oil, which solidified in about 2hours. The precipitate was collected by filtration, washed thoroughlywith water and dried in air. 22.9 g of 4-(2,5-dimethylphenoxy)-butyratewere recovered. Melting point: 63-65° C. Combustion analysis: % C, 69.21(Calc'd); 69.13 (Found); % H, 7.74 (Calc'd); 7.62 (Found); 1H NMRanalysis (d6-DMSO): δ 6.98, d, 1H; 6.71, s, 1H; 6.62, d, 1H; 3.94, t,2H; 2.40, t, 2H; 2.24, s, 3H; 2.08, s, 3H; 1.95, p, 2H.

Example 3 Preparation of 6-(3-Acetylphenoxy)hexanoic acid (Compound 2)

A round bottom flask equipped with a magnetic stirrer bar and a refluxcondenser was charged with 5.0 g (36.7 mmol) of 3-hydroxyacetophenone,8.18 g (36.7 mmol) of ethyl 6-bromohexanoate and 50 mL of ethanol. Theclear reaction mixture was treated with potassium carbonate (6.03 g,44.0 mmol) and heated to reflux. After stirring for 24 hr at reflux, thereaction mixture was cooled to 25° C., filtered through a Celite pad andconcentrated. The residue was taken up in of ethanol and treated with 15mL of 2N aqueous sodium hydroxide (30 mmol). The reaction mixture wasstirred for 6 hr at 25° C., before the ethanol was stripped off. Theresidue was acidified with aqueous 1N hydrochloric acid. The resultingsolid was isolated by filtration and purified by recrystallization fromethanol/water to yield an off-white solid, 7.57 g, mp 70-72° C.

Combustion analysis: Found: C, 66.70; H, 7.42%; C14H18O4 requires C,67.09; H, 7.25%; 1H NMR (d6-DMSO): ÿ 12.1, bs, 1H (COOH); ÿ 7.54, dt,1H, (H ortho to acetyl); ÿ 7.43, m, 2H (H's para to acetyl and OR); ÿ7.20, dd, 1H (H ortho to OR); ÿ 4.02, t, 2H, (CH2 ÿ to 0); ÿ 2.57, s, 3H(CH3 of acetyl); ÿ 2.24, t, 2H (CH2 ÿ to COOH); ÿ 1.73, p, 2H (CH2 ÿ toArO or to COOR); ÿ 1.54, p, 2H (CH2 ÿ to ArO or to COOR), ÿ 1.46, p, 2H(CH2 in middle of chain).

Example 4 Preparation of 4-(2-methylbenzyloxy)butyric acid (Compound 3)

A suspension of 2-methylphenol (16.22 g, 0.15 mol), ethyl4-bromobutanoate (33.80 g, 0.165 mol), and potassium carbonate (24.88 g,0.18 mol) in 500 mL of 2-butanone was heated at reflux under nitrogenfor 5 hours An additional 3 g of ethyl 4-bromobutanoate was added, andthe reflux was continued for an additional 25 h. The reaction wasallowed to cool to room temperature. Water and ethyl acetate were added.The organic product was extracted into the organic layer. The organiclayer was separated and washed with water and brine. It was concentratedin vacuo to yield an oil. 200 mL of water and 150 mL of a 2N aqueoussolution of NaOH was added to the oil. The mixture was stirred at roomtemperature overnight, and then heated at reflux for an hour. Themixture was cooled to room temperature and ice was added to chill themixture to about 0° C. Slow addition of a solution of 2N hydrochloricacid solution (150 ml) caused an immediate precipitation. The resultingsolid was collected by filtration, washed successively with water andwith heptane. Drying in vacuo provided 22.18 g (76%) of the titlecompound as a white solid. Mp: 79-80° C. HPLC Rt 4.96 min.; 1H NMR (DMSOd6, 300 MHz) δ: 1.97 (m, 2H), 2.15 (s, 3H), 2.42 (t, 2H), 3.97 (t 2H),6.82 (t, 1H), 6.88 (d, 1H), 7.10-7.16 (m, 2H), 12.20 (s, 1H). Anal.Calcd for C₁₁ 14 ₁₄O₃: C, 68.02; H, 7.27. Found: C, 68.04; H, 7.15.

Example 5 Preparation of 4-(3,4-dimethylphenoxy)-butyric acid (Compound4)

3,4-Dimethylphenol (24.5 g, 0.2 mol) was dissolved in 125 ml ofanhydrous ethanol. 75 ml of 21 wt % sodium ethoxide solution in ethanol(0.2 mol) was added to the solution. The mixture was stirred at roomtemperature for 10 min. Ethyl 4-bromobutyrate (40 g, 0.205 mol) was thenadded. The reaction mixture was refluxed at 80° C. for 24 h. Theprecipitate was filtered off. The filtrate was then mixed with asolution of sodium hydroxide (10 g, 0.25 mol) in 150 ml of water. Themixture was stirred at 70° C. for about 2 h, monitored with HPLC. It wasdiluted with 150 ml of water and acidified to pH 1. Oily precipitateformed immediately. It was collected by decanting the supernatant andwashed with water twice (100 ml×2). The oily precipitate solidifiedafter being kept in water at room temperature over night.

The precipitate was dissolved in 250 ml of water having pH 8-9. Anyinsoluble substance was filtered off. The filtrate was extracted withethyl acetate twice (100 ml×2). The aqueous solution was acidified topH 1. Precipitate formed immediately as oil, which solidified in about 2h. It was collected by filtration, washed thoroughly with water anddried in air. 22.9 g of 4-(2,5-dimethylphenoxy)-butyrate were recovered.Melting point: 87-89° C. Combustion analysis: % C, 69.21 (Calc'd), 69.25(Found); % H, 7.74 (Calc'd), 7.93 (Found); 1H NMR analysis (d6-DMSO): δ6.99, d, 1H; 6.72, d, 1H; 6.62, q, 1H; 3.90, t, 2H; 2.36, t, 2H; 2.16,s, 3H, 2.12, s, 3H; 1.90, p, 2H.

Example 6 Preparation of 4-(2,3-Dimethylphenoxy)butyric acid (Compound5)

2,3-Dimethylphenol (24.5 g, 0.2 mol) was dissolved in 125 ml ofanhydrous ethanol. Seventy five (75) ml of 21 wt % sodium ethoxidesolution in ethanol (0.2 mol) was added to the solution. The mixturesolution was stirred at room temperature for 10 min. Ethyl4-bromobutyrate (40 g, 0.205 mol) was then added to the solution. Thereaction mixture was refluxed at 80° C. for 24 h. The precipitate wasfiltered of. The filtrate was then mixed with a solution of sodiumhydroxide (10 g, 0.25 mol) in 150 ml of water. The mixture was stirredat 70° C. for about 2 hours, and monitored with HPLC. The mixture wasdiluted with 150 ml of water and acidified to pH 1. Oily precipitateformed immediately. The precipitate was collected by decanting thesupernatant and washed with water twice (100 ml×2). The oily precipitatesolidified after being kept in water at room temperature overnight.

The precipitate was dissolved in 250 ml of water at pH 8-9. Anyinsoluble substance was filtered off. The filtrate was extracted withethyl acetate twice (100 ml×2). The aqueous solution was acidified topH 1. Precipitate formed immediately as oil, which solidified in about 2hours. The precipitate was collected by filtration, washed thoroughlywith water and dried in air. 22.9 g of 4-(2,3-dimethylphenoxy)-butyratewere recovered. Melting point: 105-107° C. Combustion analysis (with aKF of 0.16): % C, 69.10 (Calc'd), 69.11 (Found); % H, 7.75 (Calc'd),7.79 (Found); 1H NMR analysis (d6-DMSO): δ 7.00, t, 1H; 6.74, m, 2H;3.94, t, 2H; 2.40, t, 2H; 2.20, s, 3H; 2.06, s, 3H; 1.95, p, 2H.

Example 7 Preparation of 4-(3-methylbenzyloxy)butyric acid (Compound 6)

A suspension of 3-methylphenol (200 g, 1.85 mol), ethyl 4-bromobutanoate(433 g, 2.11 mol), potassium hydroxide (155.7 g, 2.78 mol), 500 mL ofwater in 2500 mL of Dimethylsulfoxide (DMSO) in a 5-L flask was stirredat room temperature overnight. An additional 500 g water was added, andthe reaction mixture was heated at 75° C. for 2 hours. Potassiumhydroxide (155.7 g, 2.78 mol) was added. The reaction was stirred for anadditional 30 min. The thick slurry was transferred in a 22-L flask.Addition of water (6 L) to the slurry with stirring caused thedissolution of the slurry. Concentrated HCl (300 mL) was slowly added.During the reaction, the temperature of the reaction mixture wasmaintained at about 36° C. with external cooling with ice. After theaddition, the mixture was further cooled overnight to 10° C. Theresulting solid was collected by filtration, washed with 100 mL ofwater, and dried with in-house vacuum for 2 days. Further drying highvacuum overnight afforded 311.88 g of the crude material.Recryallization in a mixture of ethanol (400 mL) and water (150 mL)yielded 290.94 g (81%) of the desired product as a solid. Mp: 53-54° C.HPLC Rt 5.13 min.; 1H NMR (DMSO d6, 300 MHz) δ: 1.88 (m, 2H), 2.22 (s,3H), 2.33 (t, 2H), 3.89 (t, 2H), 6.62-6.72 (m, 3H), 7.10 (m, 1H), 12.20(s, 1H). Anal. Calcd for C11H14O3: C, 67.82; H, 7.28. Found: C, 67.90;H, 7.36.

Example 8 Preparation of 4-(3-Fluorophenoxy)butyric acid (Compound 7)

A 1 L 3-neck round bottom flask equipped with a magnetic stirrer bar anda reflux condenser was charged with 17.17 g (150 mmol) of3-fluorophenol, 33.82 g (165 mmol) of ethyl 4-bromobutyrate, 24.88 g,180 mmol potassium carbonate and 600 mL of 2-butanone. The slurry heatedto reflux. After stirring for 21 hr at reflux, the reaction mixture wascooled to 25° C., filtered and concentrated. The residue was taken up inof water and treated with 150 mL of 2N aqueous sodium hydroxide (300mmol). The reaction mixture was heated to reflux for 30 min and cooledto 25° C. The brown solution was acidified with aqueous 150 ml of 2Nhydrochloric acid. The resulting pink solid was isolated by filtrationwashing with hexanes to yield 25.04 g, mp 53-54° C. Combustion analysis:Found: C, 60.53; H, 5.79%; F, 9.84%; C₁₀H₁₁FO₃ requires C, 60.60; H,5.59%; F, 9.59%; 1H NMR (d6-DMSO): δ 12.1, bs, 1H (COOH); δ 7.20, dd,1H, (H para to OR); δ 6.7, m, 3H (aryl H's); δ 3.90, t 2H, (CH₂ α to 0);δ 2.30, t, 2H (CH₂ α to COOR); δ 1.87, p, 2H (CH₂ β to ArO and COOH).

Example 9 Preparation of 4-(2-Chlorophenoxy)butanoic acid

A 500 mL 3-neck round bottom flask equipped with a magnetic stirrer barand a reflux condenser was charged with 12.88 g (100 mmol) of2-chlorophenol, 22.54 g (110 mmol) of ethyl 4-bromobutyrate, 16.59 g,120 mmol potassium carbonate and 350 mL of 2-butanone. The slurry washeated to reflux. After stirring for 21 hr at reflux, the reactionmixture was cooled to 25° C., filtered and concentrated. The residue wastaken up in 400 mL of water and treated with 100 mL of 2N aqueous sodiumhydroxide (200 mmol). The reaction mixture was heated to reflux for 120min and cooled to 25° C. The resulting solution was acidified withaqueous 105 ml of 2N hydrochloric acid. The resulting white solid wasisolated by filtration by washing with hexanes to yield 20.68 g of4-(2-Chlorophenoxy)butanoic acid. Mp 85-87° C. Combustion analysis:Found: C, 55.97; H, 5.13%; Cl, 16.55%; C₁₀H₁₁ClO₃ requires C, 55.96; H,5.17%; Cl, 16.52%; 1H NMR (d6-DMSO): δ 12.1, bs, 1H (COOH); δ 7.42, dd,1H, (H ortho to Cl); δ 7.29, t, 1H (H para to Cl); δ 7.13, d, 1H, (Hortho to OR); δ 6.95, t, 1H (H para to OR); δ 4.07, t, 2H, (CH₂ α to O);δ 2.44, t, 2H (CH2 α to COOH); δ 1.98, p, 2H (CH2 β to ArO and COOH).

Example 10 Preparation of8-[2-(4-Chlorophenoxy)-2-methylpropionyl]-aminocaprylic acid

8-aminooctanoic acid (3.2 g, 20 mmol) and sodium hydroxide (2.0 g, 50mmol) were dissolved in 50 mL water. The solution was cooled in an icebath. 2-methyl-2-(4-chlorophenoxy)propionyl chloride (4.6 g, 20 mmol)was then added to the mixture dropwise while stirring vigorously. Themixture stirred at 25° C. for 3 hours. The cloudy basic solution wasextracted with ethyl acetate (20 ml×1) producing a clear solution. Thesolution was acidified to pH 1 with aqueous hydrochloric acid at 0° C. Asyrupy precipitate formed. The syrupy precipitate solidified after 3hours at 0° C. The resulting solid was isolated by filtration and driedin the air to afford 6.4 g of the crude product. Recrystallization fromethyl acetate/n-hexane yielded 5.9 g (84.5%) of the desired productMelting point: 94-96° C., HPLC Rt 6.42 min. Combustion analysis: Found:C, 60.88%; H, 7.42%, N, 3.87%, Cl, 9.96%; Calculated: C, 60.75%; H,7.36%; N, 3.94%; Cl, 9.96%. 1H NMR analysis (d6-DMSO): δ 12.0, broad s,1H, (COOH), δ 8.1, t 1H, (NH); δ 7.3, d, 2H (ayl H's); δ 6.85, d, 2H(aryl H's); δ 3.1, m, 2H (CH2 α NH); δ 2.20, t, 2H(CH2 α COOH); δ 1.4, m10H (rest of CH2's); δ 1.2, m, 6H (CH3's).

Example 11 Preparation of6-[2-(4-Chlorophenoxy)-2-methylpropionyl]-aminohexanoic acid

6-Aminohexanoic acid (3.0 g, 23 mmol) and sodium hydroxide (2.0 g, 50mmol) were dissolved in 50 mL water. The solution was cooled in an icebath, and 2-methyl-2-(4-chlorophenoxy)-propionyl chloride (4.6 g, 20mmol) was added to the mixture dropwise while stirring vigorously. Themixture stirred at 25° C. for 3 hours. The cloudy basic solution wasextracted with ethyl acetate (20 ml×1) producing a clear solution. Itwas acidified to pH 1 with aqueous hydrochloric acid at 0° C. A syrupyprecipitate formed. The syrupy precipitate solidified after 3 hours at0° C. The resulting solid was isolated by filtration and dried in theair. Recrystallization from ethyl acetate/n-hexane yielded 5.4 g (83.0%)of the desired product. Mp: 61-63° C. HPLC Rt 5.40 min.; Combustionanalysis: Pound: C, 58.62%; H, 6.35%; N, 4.17%; Calculated: C, 58.62%;H, 6.35%; N, 4.27%; 1H NMR analysis (d6-DMSO): δ 12.0, broad s, 1H(COOH); δ 8.1, t, 1H (NH); δ 7.3, d, 2H (aryl H); δ 6.85, d, 2H (arylH); δ 3.1, m, 2H (CH2 α NH); δ 2.15, t, 2H (CH2 α COOH); δ 1.4, s, 6H(CH3's); δ 1.4, m, 4H (CH2's); δ 1.2, m, 2H (middle CH2).

Example 12 Preparation of 4-(4-Fluoro-2-methyl-phenoxy)-butyric aid

A 200 ml 3-neck round bottom flask equipped with a magnetic stirrer barand a reflux condenser was charged with 4.42 g (34 mmol) of2-fluoro-4-methylphenol, 7.58 g (37 mmol) ethyl 4-bromobutyrate, 5.67 g(41 mmol) potassium carbonate and 130 mL of 2-butanone. After stirringfor 20 hr at reflux, the reaction mixture was cooled to 25° C. anddiluted with ethyl acetate and distilled water. The layers wereseparated. The organic layer was washed with distilled water and brine.It was dried over potassium carbonate, transferred to a 1000 ml flaskand concentrated. The precipitate was dissolved in distilled water. Thesolution was mixed with 40 ml (80 mmol) 2N aqueous sodium hydroxide. Thereaction mixture was heated to reflux for 30 min until the ester washydrolyzed (determined by HPLC). The mixture was cooled to 25° C., usingan ice bath. The amber solution was acidified with 45 ml of 2N aqueoushydrochloric acid. The resulting white solid was isolated by filtrationwashing with water twice followed by hexanes twice to yield 4.88 g (68%)of the desired product. Combustion analysis: Found: C, 62.07%; H, 6.4%;Calculated: C, 62.26%; H, 6.4%; 1H NMR (d6-DMSO): δ 12.1, s, 1H (COOH);δ 6.8-6.9, m, 3H (aryl H); δ3.8, t, 2H, (CH2 α to ArO); δ2.3, t, 2H (CH2α COOH); δ 2.1, s, 3H, (CH3 meta to F); δ1.8, m, 2H (CH2 β to COOH).

Example 13 Preparation of 4-(2-Fluoro-5-methyl-phenoxy)-butyric acid

A 200 ml 3-neck round bottom flask equipped with a magnetic stirrer barand a reflux condenser was charged with 4.99 g (40 mmol) of2-fluoro-5-methyl phenol, 9.01 g (44 mmol) of ethyl 4-bromobutyrate,6.63 g (48 mmol) potassium carbonate and 120 ml of 2-butanone. Theslurry heated to reflux. After stirring for 14.5 hr at reflux, thereaction mixture was cooled to 25° C., filtered and concentrated. Theresidue was taken up in water (200 ml) and treated with 42 ml (84 mmol)2N aqueous sodium hydroxide. The reaction mixture was heated to refluxfor 2 hours and cooled to 25° C. The yellow solution was acidified with45 ml 2N aqueous hydrochloric acid. The resulting white solid wasisolated by filtration washing with twice with water then twice withhexanes to yield 7.64 g (90%), mp 62-64° C. Combustion analysis: Found:C, 62.2%; H, 6.19%; Calculated: C, 62.26%; H, 6.17%; 1H NMR analysis(d6-DMSO): δ 12.2, broad s, 1H, (COOH); δ7.0, m, 2H (aryl H); δ 6.7, s,1H (aryl H); δ 4.0, t, 2H (CH₂ α ArO); δ 2.4, t, 2H (CH₂ α COOH); δ2.21, s, 3H (CH₃ para F); δ 1.90, m, 2H (CH₂ β COOH).

Example 14 Preparation of 5-(2-Chloro-phenoxy)-pentanoic acid

A 500 ml 3-neck round bottom flask equipped with a magnetic stirrer barand a reflux condenser was charged with 12.86 (100 mmol) 2-chlorophenol, 23.49 g (110 mmol) ethyl 5-bromovalerate, 16.60 g (120 mmol)potassium carbonate and 300 mL of 2-butanone. The slurry heated toreflux. After stirring for 20 hr at reflux (completeness confirmed byHPLC), the reaction mixture was cooled to 25° C., filtered andconcentrated. The residue was taken up in of water (300 ml) and treatedwith 100 ml (200 mmol) 2N aqueous sodium hydroxide. This suspension washeated to reflux for 2 hours until it turned to a solution. The solutionwas cooled to 25° C. Ice was added to further cool to 0° C. The yellowsolution was acidified with aqueous 105 ml 2N hydrochloric acid. Theresulting white solid was isolated by filtration washing with watertwice then hexanes twice to yield 22.24 g (97%) of the desired product,Mp 71-72° C. Combustion analysis: Found: C, 57.66%; H, 5.82%, Cl, 15.5%;Calculated: C, 57.78%; H, 5.73%; Cl, 15.5%; 1H NMR (d6-DMSO): δ 12.1,broad s, 1H (COOH); δ 7.40, dd, 1H (aryl H); δ 7.25, m, 1H (aryl H); δ7.10, dd, 1H (aryl H); δ 6.9, m, 1H (aryl A); δ 4.0, t, 2H (CH2 α ArO);δ 2.3, t, 2H (CH2 α COOH); δ 1.7, m, 4H (rest of CH2's).

Other compounds contained herein can be prepared by similar methodsknown to those skilled in the art.

Example 15 Oral Delivery of Insulin in Rats

Human recombinant insulin (ICN Biomedicals, Aurora, Ohio) was dissolvedin deionized water (ph˜6.5) to obtain stock insulin solutions having aconcentration of 15 mg/ml. Sodium salts of the delivery agent compoundsshown below in table 1 dissolved in deionized water to obtain a 200mg/ml delivery agent solution. The free acid form of delivery agent asconverted to the sodium salt by adding one equivalent of sodiumhydroxide. Solutions were vortexted, sonicated, and heated. Ifnecessary, additional sodium hydroxide was added in μl quantities toachieve uniform solubility in the delivery agent solutions. Solutionswere adjusted to a pit of 3.5-8.5 by the addition of either hydrochloricacid or sodium hydroxide, as appropriate. The insulin stock solution wasthen added to the delivery agent solutions to obtain an administrationsolution ultimately having an insulin concentration of 0.5 mg/ml. Aftersolubilization and drug addition, administration solutions were broughtto a final volume by the addition of deionized water.

Insulin was administered to male, Sprague-Dawley rats either alone or incombination with delivery agent by oral gavage (PO). Rats, weighing 0.22to 0.27 kg, were fasted for 18-24 hours prior to dosing. A Rusch 8French catheter was cut to 11 cm in length and adapted to fit a 1 mlsyringe. The syringe was filled with dosing solution and the catheterwas inserted into the at mouth and fed down the esophagus (10.0 cm). Thedosing solution was delivered by pressing the syringe plunger whileholding the rat in an upright position. The doses of delivery agent andinsulin were 200 mg/kg and 0.5 mg/g, respectively unless noted otherwisein Table 1. The dose volume was 1 ml/g.

Immediate to each blood sampling point, rats were exposed briefly (˜10seconds) to carbon dioxide until prostrate. A 77-nun capillary tube wasinserted into the retroorbital sinus. Typically, blood samples werecollected prior to dosing (time=0) and at 15, 30, 45, and 60 minutesafter dosing.

To determine the pharmacodynamic response, a hand-held glucometer(OneTouch Ultra, LifeScan—Johnson & Johnson, New Brunswick, N.J.) wasused to measure whole blood glucose after administration of insulin orinsulin and delivery agent. After discarding the first drop of blood, asmall sample (˜5-10 μls) was touched to the glucometer test strip(OneTouch Ultra, LifeScan) and a blood glucose reading was generated bythe meter. Samples at the times indicated below in Table 1 after dosing.

The following results were obtained:

TABLE 1 Oral Delivery of Insulin in Rats Average Amount of DeliveryGlucose Amount of Delivery Agent Reduction - Insulin Agent (mg/ TimeCom- (5 rats/ (mg/kg of kg of rat period pound experiment) rat weight)weight) (minutes) 1 −17.2% 0.5  200 30 1 −63.3% 0.5  200 30 1 −78.4%0.5  200 45 1  −8.8% 0.5   25 30 1 −51.4% 0.5   50 30 1 −31.4% 0.5  10030 1 −66.4% 0.5  200 45 1 −16.1% 0.05 200 15 1 −13.0% 0.10 200 30 1−34.0% 0.3  200 45 1 −45.4% 0.5  200 45 1  −0.8% 0.5   25 30 1  −5.7%0.5   50 30 1 −60.6% 0.5  100 45 1 −57.0% 0.5  200 30 1 −69.8% 0.5  20045 1 −41.5% 0.5  200 45 1 −29.7% 0.5  200 30 1 −55.5% 0.5  200 30 2−53.6% 0.5  200 45 2 −25.8% 0.5   25 45 2 −30.2% 0.5   50 45 2 −50.8%0.5  100 45 2 −18.7% 0.5  200 45 2  −1.1% 0.5  200 45 2 −13.3% 0.5  20045 2 −29.4% 0.5  200 45 2 −30.0% 0.5  200 45 2  +1.6% 0.5  200 45 2 +8.2% 0.5  200 45 2 −15.9% 0.5  200 45 2 −41.7% 0.5  200 45 3 −51.5%0.5  200 45 3 −72.8% 0.5  200 45 3 −43.2% 0.5  200 45 3 −49.1% 0.5  20045 3 −51.5% 0.5  200 45 4 −55.9% 0.5  200 45 4 −56.8% 0.5  200 45 4−55.1% 0.5  200 45 4  −2.2% 0.5  200 45 4  −6.7% 0.5  200 45 4  −8.3%0.5  200 45 4 −32.7% 0.5  200 45 4 −28.7% 0.5  200 45 5 −42.8% 0.5  20045 5 −66.3% 0.5  200 45 5 −52.3% 0.5  200 45 6 −36.0% 0.5  200 30 6−61.2% 0.5  200 30 6 −42.9% 0.5  200 45 6 −19.3% 0.5   25 30 6 −11.9%0.5   50 30 6 −47.9% 0.5  100 30 6 −14.4% 0.5  200 60 6  −3.7% 0.5   2530 6 −28.7% 0.5   50 30 6 −63.7% 0.5  100 30 6 −79.0% 0.5  200 45 6 −4.4% 0.5   25 30 6 −19.9% 0.5   50 30 6 −39.3% 0.5  100 45 6 −41.9%0.5  200 30 6 −33.9% 0.5  200 30 6 −44.7% 0.5  200 30 6 −55.3% 0.5  20030 6 −30.9% 0.25 100 45 7 −65.6   0.5  200 45 7 −68.0% 0.5  200 45 7−61.5% 0.5  200 45 7 −16.6% 0.5  200 45 8 −78.0% 0.5  200 45 8 −25.4%0.5  200 45 8 −52.0% 0.5  200 45 8 −34.0% 0.5  200 45 10   2.5% 0.5  20030 11 −63.7% 0.5  200 30 11 −78.0% 0.5  200 45 12 −78.8% 0.5  200 30 12−81.5% 0.5  200 30 12 −70.4% 0.5  200 30 12 −47.9% 0.5  100 30 12 −25.7%0.5   50 60 12 −13.1% 0.5   25 60 12 −41.0% 0.25 100 30 12 −82.1% 0.5 200 30 13 −36.5% 0.5  200 30 13 −48.4% 0.5  200 30 13 −63.7% 0.5  200 30

Example 16 Delivery of Argatroban in Rats

Argatroban was administered to rats orally with and without a deliveryagent, and intravenously without a delivery agent. The oral dose ofargatroban used was 4 mg/kg body weight. The dose of the sodium salt ofdelivery agent (4-(2,5-Dimethylphenoxy)butyric acid) was 200 mg/kg bodyweight. The ravenous dose of argatroban used was 1 mg/kg body weight

A baseline blood sample was taken from the retroorbital sinus prior todosing (time=0). Blood samples were drawn from the retroorbital sinus atvarious time points after dosing. Argatroban was administered to male,Sprague-Dawley rats either alone or in combination with delivery agentby oral gavage (PO). Typically, rats (generally weighing 0.22 to 0.27kg) were fasted for 18-24 hours prior to dosing. A Rusch $ Frenchcatheter was cut to 11 cm in length and adapted to fit a 1 ml syringe.The syringe was filled with dosing solution and the catheter wasinserted into the rat mouth and fed down the esophagus (10.0 cm). Thedosing solution was delivered by pressing the syringe plunger whileholding the rat in an upright position. The doses of delivery agent andargatroban were 200 mg/kg and 4.0 mg/kg, respectively. The dose volumewas 1 ml/kg. For intravenous dosing the dose of argatroban was 1 mg/kgand a 1 ml/kg final dose volume was also used. Immediate to each bloodsampling point, rats were exposed briefly (˜10 seconds) to carbondioxide until prostrate. A 77-mm capillary tube was inserted into theretroorbital sinus. Blood samples were collected prior to dosing(time=0) and at 10, 20, and 40 minutes after dosing.

The argatroban plasma concentrations were determined at Glaxo SmithKline via HPLC assay. The results are shown in FIGS. 1-6.

The above-mentioned patents, applications, test methods, andpublications are hereby incorporated by reference in their entirety.

Many variations of the present invention will suggest themselves tothose skilled in the art in light of the above detailed description. Allsuch obvious variations are within the fully intended scope of theappended claims.

What is claimed is:
 1. A delivery agent compound selected from the groupconsisting of:

and pharmaceutically acceptable salts thereof.
 2. The delivery agentcompound of claim 1, wherein the delivery agent is

or a pharmaceutically acceptable salt thereof.
 3. The A delivery agentcompound selected from the group consisting of claim 1, wherein thedelivery agent is

or aand pharmaceutically acceptable saltsalts thereof.
 4. The deliveryagent compound of claim 1, wherein the delivery agent is

or a pharmaceutically acceptable salt thereof.
 5. The delivery agentcompound of claim 1, wherein the delivery agent is

or a pharmaceutically acceptable salt thereof.
 6. A pharmaceuticalcomposition comprising (A) a therapeutically effective amount of abiologically active agent; (B) at least one compound selected from thegroup consisting of

and pharmaceutically acceptable salts thereof according to claim 3, and(C) a pharmaceutically acceptable excipient, wherein the biologicallyactive agent is selected from recombinant human growth hormone (rhGH),human growth hormone, bovine growth hormone, porcine growth hormone,BIBN-4096BS, growth hormone releasing factor, interferons,interleukin-1, interleukin-2, insulin, insulin-like growth factor (IGF),heparin, heparinoids, dermatans, chondroitins, calcitonin,erythropoietin (EPO), atrial naturetic factor, CPHPC, monoclonalantibodies, somatostatin, octreotide, adrenocorticotropin, gonadotropinreleasing hormone, oxytocin, leutinizing-hormone releasing-hormone,follicle stimulating hormone, glucocerebrosidase, thrombopoeitin,filgrastim, GM-CSF5, prostaglandins, cyclosporin, vasopressin, cromolynsodium, sodium chromoglycate, disodium chromoglycate, vancomycin,gallium nitrate, glucagon, DPP-4 inhibitors, peptide YY, desferoxamine(DFO), parathyroid hormone (PTH), glucagon-like peptide 1 (GLP-I)(GLP-1), argatroban; and any combination thereof.
 7. The pharmaceuticalcomposition of claim 6, wherein the biologically active agent isselected from the group consisting of insulin, leutenizing-hormonereleasing hormone, GLP-1, heparin, recombinant human growth hormone,argatroban, and any combination thereof.
 8. The pharmaceuticalcomposition of claim 6, wherein the biologically active agent isargatroban.
 9. The pharmaceutical composition of claim 6, wherein thebiologically active agent is insulin.
 10. The pharmaceutical compositionof claim 6, wherein the biologically active agent is human growthhormone.
 11. The pharmaceutical composition of claim 6, wherein thegrowth hormone biologically active agent is human growth hormone,recombinant human growth hormone (rhGH), bovine growth hormone orporcine growth hormone.
 12. The pharmaceutical composition of claim 6,wherein the calcitonin is salmon calcitonin, eel calcitonin or humancalcitonin.
 13. The pharmaceutical composition of claim 6, wherein theinsulin is porcine insulin, bovine insulin, human insulin or humanrecombinant insulin.
 14. The pharmaceutical composition of claim 6,wherein the heparin is unfractionated heparin, low molecular weightheparin, very low molecular weight heparin or ultra low molecular weightheparin.
 15. The pharmaceutical composition of claim 6, wherein theinsulin-like growth factor is IGF-I.
 16. A dosage unit form comprising apharmaceutical composition of claim
 6. 17. The dosage unit form of claim16, wherein the dosage unit form is a tablet or capsule.
 18. Apharmaceutical composition comprising (A) a therapeutically effectiveamount of a biologically active agent;

and pharmaceutically acceptable salts thereof; and (C) apharmaceutically acceptable excipient; wherein the biologically activeagent is selected from calcitonin, insulin, GLP-1, heparin, argatroban,and any combination thereof.
 19. A pharmaceutical composition comprising(A) a therapeutically effective amount of a biologically active agent;(B) a compound selected from:

and pharmaceutically acceptable salts thereof; and (C) apharmaceutically acceptable excipient; wherein the biologically activeagent is selected from calcitonin, insulin, GLP-1, heparin, argatroban;and any combination thereof.
 20. A pharmaceutical composition comprising(A) a therapeutically effective amount of a biologically active agent;(B) a compound selected from:

and pharmaceutically acceptable salts thereof; and (C) apharmaceutically acceptable excipient; wherein the biologically activeagent is selected from BIBN-4096BS, growth hormone releasing factor,interferons, interleukin-1, interleukin-2, insulin, insulin-like growthfactor (IGF), heparin, heparinoids, dermatans, chondroitins, calcitonin,erythropoietin (EPO), atrial naturetic factor, CPHPC, monoclonalantibodies, somatostatin, octreotide, adrenocorticotropin, gonadotropinreleasing hormone, oxytocin, leutinizing-hormone releasing-hormone,follicle stimulating hormone, glucocerebrosidase, thrombopoeitin,filgrastim, GM-CSF5, prostaglandins, cyclosporin, vasopressin, cromolynsodium, sodium chromoglycate, disodium chromoglycate, vancomycin,gallium nitrate, glucagon, DPP-4 inhibitors, peptide YY, desferoxamine(DFO), parathyroid hormone (PTH), glucagon-like peptide 1 (GLP-I)(GLP-1), argatroban; and any combination thereof.
 21. A pharmaceuticalcomposition comprising (A) a therapeutically effective amount of abiologically active agent; (B) a compound selected from:

and pharmaceutically acceptable salts thereof; and (C) apharmaceutically acceptable excipient; wherein the biologically activeagent is selected from BIBN-4096BS, growth hormone releasing factor,interferons, interleukin-1, interleukin-2, insulin, insulin-like growthfactor (IGF), heparin, heparinoids, dermatans, chondroitins, calcitonin,erythropoietin (EPO), atrial naturetic factor, CPHPC, monoclonalantibodies, somatostatin, octreotide, adrenocorticotropin, gonadotropinreleasing hormone, oxytocin, leutinizing-hormone releasing-hormone,follicle stimulating hormone, glucocerebrosidase, thrombopoeitin,filgrastim, GM-CSF5, prostaglandins, cyclosporin, vasopressin, cromolynsodium, sodium chromoglycate, disodium chromoglycate, vancomycin,gallium nitrate, glucagon, DPP-4 inhibitors, peptide YY, desferoxamine(DFO), parathyroid hormone (PTH), glucagon-like peptide 1 (GLP-I)(GLP-1), argatroban; and any combination thereof.
 22. The compositionaccording to claim 20, wherein the biologically active agent is selectedfrom calcitonin, insulin, PTH, GLP-1, heparin, argatroban; and anycombination thereof.
 23. The composition according to claim 21, whereinthe biologically active agent is selected from calcitonin, insulin, PTH,GLP-1, heparin, argatroban; and any combination thereof.
 24. A methodfor administering a biologically active agent comprising orallyadministering to a human a pharmaceutical composition comprising: (A) atherapeutically effective amount of a biologically active agent whereinthe biologically active agent is a polypeptide; (B) a compound selectedfrom

and pharmaceutically acceptable salts thereof; and (C) apharmaceutically acceptable excipient.
 25. The method of claim 24,wherein compound (B) is selected from

and pharmaceutically acceptable salts thereof.
 26. The method of claim24, wherein compound (B) is selected from

and pharmaceutically acceptable salts thereof.
 27. The method of claim24, wherein the polypeptide is GLP-1.
 28. The method of claim 25,wherein the polypeptide is GLP-1.
 29. The method of claim 26, whereinthe polypeptide is GLP-1.
 30. The composition of claim 18, wherein thebiologically active agent is insulin, heparin, or any combinationthereof.
 31. The composition of claim 19, wherein the biologicallyactive agent is insulin, heparin, or any combination thereof.
 32. Themethod of claim 24, wherein the biologically active agent is insulin,peptide YY, human growth hormone, heparin, luteinizing hormone-releasinghormone, or any combination thereof.